Therapeutic Effect And Mechanism Of Splenectomy Against Liver Fibrosis In Mice Induced By ConA Through Promoting The Polarization Of MDSCs

Objective: To investigate the therapeutic effects of splenectomy on the development of liver fibrosis in concanavalin A-induced immunologic liver injury and the underlying mechanism.Methods: Eight-week-old female Balb/c mice were randomly divided into six groups:control group,Con A model group,Con A+dexamethasone group,Con A+sham-operation group,Con A+splenectomy group and Con A+ splenectomy +dexamethasone group.Con A?12.5mg/kg?was injected through tail vein into female Balb/c mice once a week.After 5 weeks,splenectomy and sham-operation were performed,while dexamethasone was injected into mice of Con A + dexamethasone group and Con A + dexamethasone + splenectomy group once every other day.The injection of Con A was continued up to 7 weeks.Liver damage of mice with hepatitis was evaluated with respect to serum levels of alanine transaminase?ALT?,aspartate aminotransferase?AST?and hematoxylin and eosin?HE?.The expression level of?-SMA was used to evaluate the degree of activation of hepatic stellate cells.The liver fibrosis was evaluated by Masson staining and Points-scoring system of Ishak.The polarization of MDSCs and monocytes were detected by flow cytometric analysis.The expression of transcription factor of NF-kappa B P50 and NF-kappa B P65 in liver of mice were detected by western blot.The expressions of M1 and M2 macrophages in mouse liver were detected by Real-time PCR and immunofluorescence.Results: In the fifth week,ConA model group showed higher serum levels of ALT?p<0.05?and AST?p<0.05?and severe liver inflammation by H&E than control group.The level of ?-SMA was significantly higher than that of normal control group?p<0.05?and mild liver fibrosis was detected by Masson staining.In seventh weeks,compared with Con A model group,Con A + dexamethasone group,Con A +splenectomy group and Con A + dexamethasone + splenectomy group showed significantly lower serum levels of ALT?p<0.05?,AST?p<0.05?and ?-SMA and less severe liver inflammation by H&E.After treatment with Con A for 7 weeks,remarkable liver fibrosis was detected by Masson staining,while liver fibrosis was significantly reduced in Con A + dexamethasone group,Con A + splenectomy group and Con A + dexamethasone + splenectomy group.The proportion of CD11b⁺Ly6C^highMDSCs was significantly lower in Con A model group than that of control group?p<0.05?,while it was higher in Con A+dexamethasone group,Con A+splenectomy group and Con A + dexamethasone+ splenectomy group?p<0.05?.The ratio of monocyte was higher in Con A model group than that of control group?p<0.05?,while it was significantly lower in Con A + dexamethasone group,Con A +splenectomy group and Con A+ dexamethasone + splenectomy group?p<0.05?.The proportion of F4/80+CD206+ monocyte was lower in Con A model group than that in Con A + dexamethasone group and Con A + splenectomy group.The level of NF-k B p65 was significantly lower while the level of NF-k B p50 was significantly higher in Con A + dexamethasone group,Con A + splenectomy group and Con A +dexamethasone + splenectomy group than those of Con A group?p<0.05?.The ratio of macrophages and M1 macrophages were significantly higher in Con A model group than control group?p<0.05?,while it was significantly lower in Con A +dexamethasone group,Con A + splenectomy group and Con A + dexamethasone +splenectomy group?p<0.05?.The level of ARG-1,IL-4 and IL-10 were increased in Con A+ dexamethasone group,Con A + splenectomy group and Con A +dexamethasone + splenectomy group than Con A model group?p<0.05?.Conclusion: Con A can be used to establish a model of chronic immune liver fibrosis in Balb/c mice after continuous injection for 5 weeks.Splenectomy could inhibit the activation of hepatic stellate cells and delay the progression of hepatic fibrosis in mice with immunological liver injury induced by Con A.In Con A-induced immunologic liver injury,splenectomy can promote the polarization of CD11b⁺Ly6C^highMDSCs,further promote the differentiation of M2 macrophages through increasing the expression of P50 homodimers,which can slow down the progression of immunologic liver injury induced by Con A into liver fibrosis.